INTERNATIONAL STANDARD ISO 11285 IDF 175 First edition 2004-09-01 Milk Determination of lactulose content Enzymatic method Lait Détermination de la teneur en lactulose Méthode enzymatique Reference numbers ISO 11285:2004(E) ISO and IDF 2004
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Foreword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member bodies). The work of preparing International Standards is normally carried out through ISO technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization. International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2. The main task of technical committees is to prepare International Standards. Draft International Standards adopted by the technical committees are circulated to the member bodies for voting. Publication as an International Standard requires approval by at least 75 % of the member bodies casting a vote. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. ISO 11285 IDF 175 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 5, Milk and milk products, and the International Dairy Federation (IDF), in collaboration with AOAC International. It is being published jointly by ISO and IDF and separately by AOAC International. ISO and IDF 2004 All rights reserved iii
Foreword IDF (the International Dairy Federation) is a worldwide federation of the dairy sector with a National Committee in every member country. Every National Committee has the right to be represented on the IDF Standing Committees carrying out the technical work. IDF collaborates with ISO and AOAC International in the development of standard methods of analysis and sampling for milk and milk products. Draft International Standards adopted by the Action Teams and Standing Committees are circulated to the National Committees for voting. Publication as an International Standard requires approval by at least 50 % of IDF National Committees casting a vote. ISO 11285 IDF 175 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 5, Milk and milk products, and the International Dairy Federation (IDF), in collaboration with AOAC International. It is being published jointly by ISO and IDF and separately by AOAC International. All work was carried out by the Joint ISO/IDF/AOAC Action Team, Characterization of milk and milk products according to heat treatment, of the Standing Committee on Minor components and characterization of physical properties, under the aegis of its project leader, Mrs E. Lechner (DE). This edition of ISO 11285 IDF 175 cancels and replaces IDF 175:1995, which has been technically revised. iv ISO and IDF 2004 All rights reserved
INTERNATIONAL STANDARD ISO 11285:2004(E) Milk Determination of lactulose content Enzymatic method 1 Scope This International Standard specifies an enzymatic method for the determination of the lactulose content of milk. 2 Terms and definitions For the purposes of this document, the following terms and definitions apply. 2.1 lactulose content mass of substances determined by the procedure specified in this International Standard NOTE The lactulose content is expressed as milligrams per kilogram. 3 Principle Fat and protein are precipitated by the addition of zinc sulfate and potassium hexacyanoferrate(ii) solution and are then removed by filtration. Lactose and lactulose are hydrolysed to galactose and glucose, or galactose and fructose, respectively, in the presence of the enzyme β-d-galactosidase (β-gal). The amount of liberated fructose is stoichiometric with the amount of lactulose: β-gal lactose + H 2 O galactose + glucose (1) β-gal lactulose + H 2 O galactose + fructose (2) Lactose is present in milk in significantly higher amounts than lactulose. Thus, the glucose present after hydrolysis would interfere with the determination of fructose. Therefore, the glucose is mostly oxidized by glucose oxidase (GOD) to gluconic acid in the presence of oxygen: GOD glucose + H 2 O + O 2 gluconic acid + H 2 O 2 (3) By this means it is possible to determine small amounts of lactulose in the presence of excess lactose. The hydrogen peroxide formed in reaction (3) is destroyed by catalase. This reaction is used to provide the oxygen which is required for the oxidation of the glucose: catalase 2 H 2 O 2 2 H 2 O + O 2 (4) The remaining unoxidized glucose and the fructose produced by the hydrolysis of the lactulose are phosphorylated by means of adenosine triphosphate (ATP) in the presence of hexokinase (HK) to yield glucose-6-phosphate and fructose-6-phosphate, respectively. ISO and IDF 2004 All rights reserved 1